Publications by authors named "Maria Rosaria Cera"

9 Publications

  • Page 1 of 1

High-throughput screening identifies histone deacetylase inhibitors that modulate GTF2I expression in 7q11.23 microduplication autism spectrum disorder patient-derived cortical neurons.

Mol Autism 2020 11 19;11(1):88. Epub 2020 Nov 19.

High Definition Disease Modelling Lab: Stem Cell and Organoid Epigenetics, IEO, European Institute of Oncology IRCCS, Via Adamello 16, 20139, Milan, Italy.

Background: Autism spectrum disorder (ASD) is a highly prevalent neurodevelopmental condition affecting almost 1% of children, and represents a major unmet medical need with no effective drug treatment available. Duplication at 7q11.23 (7Dup), encompassing 26-28 genes, is one of the best characterized ASD-causing copy number variations and offers unique translational opportunities, because the hemideletion of the same interval causes Williams-Beuren syndrome (WBS), a condition defined by hypersociability and language strengths, thereby providing a unique reference to validate treatments for the ASD symptoms. In the above-indicated interval at 7q11.23, defined as WBS critical region, several genes, such as GTF2I, BAZ1B, CLIP2 and EIF4H, emerged as critical for their role in the pathogenesis of WBS and 7Dup both from mouse models and human studies.

Methods: We performed a high-throughput screening of 1478 compounds, including central nervous system agents, epigenetic modulators and experimental substances, on patient-derived cortical glutamatergic neurons differentiated from our cohort of induced pluripotent stem cell lines (iPSCs), monitoring the transcriptional modulation of WBS interval genes, with a special focus on GTF2I, in light of its overriding pathogenic role. The hits identified were validated by measuring gene expression by qRT-PCR and the results were confirmed by western blotting.

Results: We identified and selected three histone deacetylase inhibitors (HDACi) that decreased the abnormal expression level of GTF2I in 7Dup cortical glutamatergic neurons differentiated from four genetically different iPSC lines. We confirmed this effect also at the protein level.

Limitations: In this study, we did not address the molecular mechanisms whereby HDAC inhibitors act on GTF2I. The lead compounds identified will now need to be advanced to further testing in additional models, including patient-derived brain organoids and mouse models recapitulating the gene imbalances of the 7q11.23 microduplication, in order to validate their efficacy in rescuing phenotypes across multiple functional layers within a translational pipeline towards clinical use.

Conclusions: These results represent a unique opportunity for the development of a specific class of compounds for treating 7Dup and other forms of intellectual disability and autism.
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http://dx.doi.org/10.1186/s13229-020-00387-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7677843PMC
November 2020

Discovery of Reversible Inhibitors of KDM1A Efficacious in Acute Myeloid Leukemia Models.

ACS Med Chem Lett 2020 May 13;11(5):754-759. Epub 2020 Feb 13.

Department of Experimental Oncology, Academic Drug Discovery, European Institute of Oncology IRCCS, Via Adamello 16, 20139 Milan, Italy.

Lysine-specific demethylase 1 (LSD1 or KDM1A) is a FAD-dependent enzyme that acts as a transcription corepressor or coactivator by regulating the methylation status of histone H3 lysines K4 and K9, respectively. KDM1A represents an attractive target for cancer therapy. While, in the past, the main medicinal chemistry strategy toward KDM1A inhibition was based on the optimization of ligands that irreversibly bind the FAD cofactor within the enzyme catalytic site, we and others have also identified reversible inhibitors. Herein we reported the discovery of 5-imidazolylthieno[3,2-]pyrroles, a new series of KDM1A inhibitors endowed with picomolar inhibitory potency, active in cells and efficacious after oral administration in murine leukemia models.
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http://dx.doi.org/10.1021/acsmedchemlett.9b00604DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7236255PMC
May 2020

Novel potent inhibitors of the histone demethylase KDM1A (LSD1), orally active in a murine promyelocitic leukemia model.

Future Med Chem 2017 07 19;9(11):1161-1174. Epub 2017 Jul 19.

Department of Experimental Oncology, Academic Drug Discovery, European Institute of Oncology, Ifom-IEO-Campus, via Adamello 16, 20139 Milan, Italy.

Background: Histone lysine demethylases (KDMs) are well-recognized targets in oncology drug discovery. They function at the post-translation level controlling chromatin conformation and gene transcription. KDM1A is a flavin adenine dinucleotide-dependent amine oxidase, overexpressed in several tumor types, including acute myeloid leukemia, neuroblastoma and non-small-cell lung cancer. Among the many known monoamine oxidase inhibitors screened for KDM1A inhibition, tranylcypromine emerged as a moderately active hit, which irreversibly binds to the flavin adenine dinucleotide cofactor.

Material & Methods: The KDM1A inhibitors 5a-w were synthesized and tested in vitro and in vivo. The biochemical potency was determined, modulation of target in cells was demonstrated on KDM1A-dependent genes and the anti-clonogenic activity was performed in murine acute promyelocytic Leukemia (APL) blasts. An in vivo efficacy experiment was conducted using an established murine promyelocytic leukemia model.

Results: We report a new series of tranylcypromine derivatives substituted on the cyclopropyl moiety, endowed with high potency in both biochemical and cellular assays.

Conclusion: The most interesting derivative (5a) significantly improved survival rate after oral administration in a murine model of promyelocitic leukemia.
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http://dx.doi.org/10.4155/fmc-2017-0003DOI Listing
July 2017

Evading Pgp activity in drug-resistant cancer cells: a structural and functional study of antitubulin furan metotica compounds.

Mol Cancer Ther 2012 May 21;11(5):1103-11. Epub 2012 Mar 21.

Target Structure-Based Drug Discovery Group, SAIC-Frederick, Inc., Frederick, Maryland, USA.

Tumor resistance to antitubulin drugs resulting from P-glycoprotein (Pgp) drug-efflux activity, increased expression of the βIII tubulin isotype, and alterations in the drug-binding sites are major obstacles in cancer therapy. Consequently, novel antitubulin drugs that overcome these challenges are of substantial interest. Here, we study a novel chemotype named furan metotica that localizes to the colchicine-binding site in β-tubulin, inhibits tubulin polymerization, and is not antagonized by Pgp. To elucidate the structure-activity properties of this chiral chemotype, the enantiomers of its most potent member were separated and their absolute configurations determined by X-ray crystallography. Both isomers were active and inhibited all 60 primary cancer cell lines tested at the U.S. National Cancer Institute. They also efficiently killed drug-resistant cancer cells that overexpressed the Pgp drug-efflux pump 10(6)-fold. In vitro, the R-isomer inhibited tubulin polymerization at least 4-fold more potently than the S-isomer, whereas in human cells the difference was 30-fold. Molecular modeling showed that the two isomers bind to β-tubulin in distinct manners: the R-isomer binds in a colchicine-like mode and the S-isomer in a podophyllotoxin-like fashion. In addition, the dynamic binding trajectory and occupancy state of the R-isomer were energetically more favorable then those of the S-isomer, explaining the observed differences in biologic activities. The ability of a racemic drug to assume the binding modes of two prototypical colchicine-site binders represents a novel mechanistic basis for antitubulin activity and paves the way toward a comprehensive design of novel anticancer agents.
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http://dx.doi.org/10.1158/1535-7163.MCT-11-1018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3349764PMC
May 2012

The synaptic proteins neurexins and neuroligins are widely expressed in the vascular system and contribute to its functions.

Proc Natl Acad Sci U S A 2009 Dec 19;106(49):20782-7. Epub 2009 Nov 19.

Department of Oncological Sciences and Institute for Cancer Research and Treatment, University of Torino School of Medicine, 10060 Candiolo (TO), Italy.

Unlike other neuronal counterparts, primary synaptic proteins are not known to be involved in vascular physiology. Here, we demonstrate that neurexins and neuroligins, which constitute large and complex families of fundamental players in synaptic activity, are produced and processed by endothelial and vascular smooth muscle cells throughout the vasculature. Moreover, they are dynamically regulated during vessel remodeling and form endogenous complexes in large vessels as well as in the brain. We used the chicken chorioallantoic membrane as a system to pursue functional studies and demonstrate that a monoclonal recombinant antibody against beta-neurexin inhibits angiogenesis, whereas exogenous neuroligin has a role in promoting angiogenesis. Finally, as an insight into the mechanism of action of beta-neurexin, we show that the anti-beta-neurexin antibody influences vessel tone in isolated chicken arteries. Our finding strongly supports the idea that even the most complex and plastic events taking place in the nervous system (i.e., synaptic activity) share molecular cues with the vascular system.
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http://dx.doi.org/10.1073/pnas.0809510106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2791601PMC
December 2009

JAM-A promotes neutrophil chemotaxis by controlling integrin internalization and recycling.

J Cell Sci 2009 Jan;122(Pt 2):268-77

FIRC Institute of Molecular Oncology, Milan, Italy.

The membrane-associated adhesion molecule JAM-A is required for neutrophil infiltration in inflammatory or ischemic tissues. JAM-A expressed in both endothelial cells and neutrophils has such a role, but the mechanism of action remains elusive. Here we show that JAM-A has a cell-autonomous role in neutrophil chemotaxis both in vivo and in vitro, which is independent of the interaction of neutrophils with endothelial cells. On activated neutrophils, JAM-A concentrates in a polarized fashion at the leading edge and uropod. Surprisingly, a significant amount of this protein is internalized in intracellular endosomal-like vesicles where it codistributes with integrin beta1. Clustering of beta1 integrin leads to JAM-A co-clustering, whereas clustering of JAM-A does not induce integrin association. Neutrophils derived from JAM-A-null mice are unable to correctly internalize beta1 integrins upon chemotactic stimuli and this causes impaired uropod retraction and cell motility. Consistently, inhibition of integrin internalization upon treatment with BAPTA-AM induces a comparable phenotype. These data indicate that JAM-A is required for the correct internalization and recycling of integrins during cell migration and might explain why, in its absence, the directional migration of neutrophils towards an inflammatory stimulus is markedly impaired.
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http://dx.doi.org/10.1242/jcs.037127DOI Listing
January 2009

Unique role of junctional adhesion molecule-a in maintaining mucosal homeostasis in inflammatory bowel disease.

Gastroenterology 2008 Jul 11;135(1):173-84. Epub 2008 Apr 11.

Division of Gastroenterology, Istituto di Ricerca e Cura a Carattere Scientifico Istituto Clinico Humanitas, University of Milan, Rozzano, Italy.

Background & Aims: Junctional adhesion molecule-A (JAM-A) is localized at the tight junctions and controls leukocyte migration into the tissues. However, its functional role in inflammatory bowel disease (IBD) is unexplored.

Methods: Control, Crohn's disease (CD), and ulcerative colitis (UC) tissue specimens were studied for JAM-A expression, as well as the colon of mice given dextran sodium sulfate (DSS). Wild-type and JAM-A(-/-), Tie-2-Cre-JAM-A(-/-) (endothelial/hematopoietic-specific JAM inactivation) mice were studied for susceptibility to DSS. Disease activity and colonic inflammation were assessed using a disease activity index histology and endoscopy, and mucosal cytokines were measured by enzyme-linked immunosorbent assay. JAM-A function was investigated by RNA silencing in epithelial cells, and apoptosis was measured.

Results: In both CD and UC, as well as in experimental colitis, there is a loss of epithelial but not endothelial JAM-A expression. Deletion of JAM-A results in a dramatic increase in susceptibility to DSS colitis, as assessed by weight loss, disease activity index, histologic and endoscopic severity, and strikingly high mortality rates. This is not caused by the absence of JAM-A in the endothelial or hematopoietic compartments because Tie-2-Cre-JAM-A(-/-) mice are no more susceptible to DSS colitis than wild-type animals. JAM-A(-/-) mice displayed increased intestinal permeability and inflammatory cytokine production, and marked epithelial apoptosis. Silencing of JAM-A in intestinal epithelial cells resulted in increased permeability in vitro.

Conclusions: Our results show a nonredundant and novel role of JAM-A in controlling mucosal homeostasis by regulating the integrity and permeability of epithelial barrier function.
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http://dx.doi.org/10.1053/j.gastro.2008.04.002DOI Listing
July 2008

Junctional adhesion molecule-A-deficient polymorphonuclear cells show reduced diapedesis in peritonitis and heart ischemia-reperfusion injury.

Proc Natl Acad Sci U S A 2005 Jul 18;102(30):10634-9. Epub 2005 Jul 18.

Fondazione Italiana per la Ricerca sul Cancro, Institute of Molecular Oncology, 20139 Milan, Italy.

Junctional Adhesion Molecule-A (JAM-A) is a transmembrane adhesive protein expressed at endothelial junctions and in leukocytes. Here we report that JAM-A is required for the correct infiltration of polymorphonuclear leukocytes (PMN) into an inflamed peritoneum or in the heart upon ischemia-reperfusion injury. The defect was not observed in mice with an endothelium-restricted deficiency of the protein but was still detectable in mice transplanted with bone marrow from JAM-A(-/-) donors. Microscopic examination of mesenteric and heart microvasculature of JAM-A(-/-) mice showed high numbers of PMN adherent on the endothelium or entrapped between endothelial cells and the basement membrane. In vitro, in the absence of JAM-A, PMN adhered more efficiently to endothelial cells and basement membrane proteins, and their polarized movement was strongly reduced. This paper describes a nonredundant role of JAM-A in controlling PMN diapedesis through the vessel wall.
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http://dx.doi.org/10.1073/pnas.0500147102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1180753PMC
July 2005

Increased DC trafficking to lymph nodes and contact hypersensitivity in junctional adhesion molecule-A-deficient mice.

J Clin Invest 2004 Sep;114(5):729-38

Department of Vascular Biology, Italian Foundation for Cancer Research (FIRC) Institute of Molecular Oncology, Milan, Italy.

Junctional adhesion molecule-A (JAM-A) is a transmembrane adhesive protein expressed at endothelial junctions and in leukocytes. In the present work, we found that DCs also express JAM-A. To evaluate the biological relevance of this observation, Jam-A(-/-) mice were generated and the functional behavior of DCs in vitro and in vivo was studied. In vitro, Jam-A(-/-) DCs showed a selective increase in random motility and in the capacity to transmigrate across lymphatic endothelial cells. In vivo, Jam-A(-/-) mice showed enhanced DC migration to lymph nodes, which was not observed in mice with endothelium-restricted deficiency of the protein. Furthermore, increased DC migration to lymph nodes was associated with enhanced contact hypersensitivity (CHS). Adoptive transfer experiments showed that JAM-A-deficient DCs elicited increased CHS in Jam-A(+/+) mice, further supporting the concept of a DC-specific effect. Thus, we identified here a novel, non-redundant role of JAM-A in controlling DC motility, trafficking to lymph nodes, and activation of specific immunity.
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http://dx.doi.org/10.1172/JCI21231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC514585PMC
September 2004